STUDY MATERIAL

Q.1 Write detailed account of c₃ pathway?

All photosynthetic eukaryotes, from the most primitive alga to the most advanced angiosperm, reduce CO2 to carbohydrate

via the same basic mechanism: the photosynthetic carbon reduction cycle originally described for C3 species (the Calvin cycle, or reductive pentose phosphate [RPP] cycle). Other metabolic pathways associated with the photosynthetic fixation of CO2, such as the C4 photosynthetic carbon assimilation cycle and the photorespiratory carbon oxidation cycle, are either auxiliary to or dependent on the basic Calvin cycle. In this section we will examine how CO2 is fixed by the

Calvin cycle through the use of ATP and NADPH generated by the light reactions (Figure 8.1), and how the Calvin cycle

is regulated.

The Calvin Cycle Has Three Stages: Carboxylation, Reduction, and Regeneration

The Calvin cycle was elucidated as a result of a series of elegant experiments by Melvin Calvin and his colleagues in the 1950s, for which a Nobel Prize was awarded in 1961. the Calvin cycle, CO2 and water from the environment are enzymatically combined with a five-carbon acceptor molecule to generate two molecules of a three-carbon intermediate. This intermediate (3-phosphoglycerate)is reduced to carbohydrate by use of the ATP and NADPH generated photochemically. The cycle is completed by regeneration of the five-carbon acceptor (ribulose-1,5-bisphosphate, abbreviated RuBP).

The Calvin cycle proceeds in three stages :

1. Carboxylation of the CO2 acceptor ribulose-1,5-bisphosphate, forming two molecules of 3-phosphoglycerate, the first stable intermediate of the Calvin cycle

2. Reduction of 3-phosphoglycerate, forming gyceraldehyde- 3-phosphate, a carbohydrate

3. Regeneration of the CO2 acceptor ribulose-1,5-bisphosphate from glyceraldehyde-3-phosphate

The carbon in CO2 is the most oxidized form found in nature (+4). The carbon of the first stable intermediate, 3- phosphoglycerate, is more reduced (+3), and it is further reduced in the glyceraldehyde-3-phosphate product (+1). Overall, the early reactions of the Calvin cycle complete the reduction of atmospheric carbon and, in so doing, facilitate its incorporation into organic compounds.

The Carboxylation of Ribulose Bisphosphate Is Catalyzed by the Enzyme Rubisco

CO₂ enters the Calvin cycle by reacting with ribulose-1,5-bisphosphate to yield two molecules of 3-phosphoglycerate  a reaction catalyzed by the chloroplast enzyme ribulose bisphosphate carboxylase/oxygenase, referred to as rubisco

CO₂ is transformed to 3-phosphoglycerate, the first stable intermediate organic molecule with three carbon atoms, in this metabolic cycle supplement. The C3 carbon fixation pathway is the Calvin cycle's first step.

Calvin cycle is also known as the C3 cycle or light-independent or dark reaction of photosynthesis. However, it is most active during the day when NADPH and ATP are abundant. To build organic molecules, the plant cells use raw materials provided by the light reactions:

1. Energy: ATP provided by cyclic and noncyclic photophosphorylation, which drives the endergonic reactions.

2. Reducing power: NADPH provided by photosystem I is the source of hydrogen and the energetic electrons required to bind them to carbon atoms. Much of the light energy captured during photosynthesis ends up in the energy-rich C—H bonds of sugars.Plants store light energy in the form of carbohydrates, primarily starch and sucrose. The carbon and oxygen required for this process are obtained from CO2, and the energy for carbon fixation is derived from the ATP and NADPH produced during the photosynthesis process. The conversion of CO2 to carbohydrate is called Calvin Cycle or C3 cycle and is named after Melvin Calvin who discovered it. The plants that undergo the Calvin cycle for carbon fixation are known as C3 plants.Calvin Cycle requires the enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase commonly called RuBisCO. It generates the triose phosphates, 3-phosphoglycerate (3-PGA), glyceraldehyde-3P (GAP), and dihydroxyacetone phosphate (DHAP), all of which are used to synthesize the hexose phosphates fructose-1,6-bisphosphate and fructose 6-phosphate.

Stages of C3 Cycle

Calvin cycle or C3 cycle can be divided into three main stages:

Carbon fixation

The key step in the Calvin cycle is the event that reduces CO2. CO2 binds to RuBP in the key process called carbon fixation, forming two-three carbon molecules of phosphoglycerate. The enzyme that carries out this reaction is Ribulose bisphosphate carboxylase/oxygenase, which is very large with a four-subunit and present in the chloroplast stroma. This enzyme works very sluggishly, processing only about three molecules of RuBP per second (a typical enzyme process of about 1000 substrate molecules per second). In a typical leaf, over 50% of all the protein is RuBisCO. It is thought to be the most abundant protein on the earth.

Reduction

It is the second stage of Calvin cycle. The 3-PGA molecules created through carbon fixation are converted into molecules of simple sugar – glucose.              This stage obtains energy from ATP and NADPH formed during the light-dependent reactions of photosynthesis. In this way, Calvin cycle becomes a pathway in which plants convert sunlight energy into long-term storage molecules, such as sugars. The energy from the ATP and NADPH is transferred to the sugars.This step is known as reduction since electrons are transferred to 3-PGA molecules to form glyceraldehyde-3 phosphate.

Regeneration

It is the third stage of the Calvin cycle and is a complex process that requires ATP. In this stage, some of the G3P molecules are used to produce   glucose, while others are recycled to regenerate the RuBP acceptor.

Products of C3 Cycle

• One molecule of carbon is fixed at each turn of the Calvin cycle.
• One molecule of glyceraldehyde-3 phosphate is created in three turns of the Calvin cycle.
• Two molecules of glyceraldehyde-3 phosphate combine together to form one glucose molecule.
• 3 ATP and 2 NADPH molecules are used during the reduction of 3-phosphoglyceric acid to glyceraldehyde-3 phosphate and in the regeneration of RuBP.
• 18 ATP and 12 NADPH are consumed in the production of 1 glucose molecule.

Q.2 Write on RUBISCO?

RUBISCO is Ribulose-1,5-bisphosphate carboxylase/oxygenase. RUBISCO catalyses the rate-limiting move within the Calvin-Benson cycle, which converts atmospheric carbon into biologically useful carbon. Rubisco’s slow catalytic rate, as well as relatively low specificity, necessarily requires the production of large amounts of this enzyme. To construct a more efficient Rubisco plant, we must first understand better the folding as well as assembly process. The most abundant enzyme on Earth is ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), and autotrophic organisms use it to transform CO2 into organic molecules through the Calvin-Benson route (Andersson and Backlund, 2008). Rubisco catalyses photosynthesizing decarbonisation as well as photorespiratory carbon oxidation in the presence of its substrate’s ribulose-1,5-bisphosphate and CO2 or O2. Due to the sheer bad catalytic characteristic of Rubisco CO2 fixation, this enzyme must be abundant. Ribulose bisphosphate Carboxylase-Oxygenase or RuBisCO is the most abundant protein in the biosphere. It catalyses the first step of carbon fixation in the Calvin cycle during photosynthesis. It is the common pathway of carbon fixation in all plants, i.e. C₃, C₄ and CAM plants. RuBisCO catalyses the carboxylation of ribulose bisphosphate (RuBP), which is the primary acceptor of CO₂ in the C₃ pathway or the Calvin cycle. It is the first step of the Calvin cycle. RuBisCO is present in the mesophyll cells of C₃ plants and bundle sheath cells of C₄ plants. Ribulose bisphosphate (RuBP) is a 5-carbon compound. It acts as a primary carbon dioxide acceptor in the Calvin cycle, and is converted into 2 molecules of 3-PGA (3-phosphoglyceric acid) by the action of RuBisCO. RuBisCO, as the name suggests, has both carboxylase and oxygenase activity. When molecular oxygen is the substrate, it converts RuBP to one molecule of phosphoglycerate and phosphoglycolate in the process called photorespiration.

Role of RuBisCO in Photosynthesis

RuBisCO catalyses the first step of carbon fixation in the Calvin cycle. Calvin cycle occurs in all plants, i.e. C₃, C₄ and CAM.

The first step of the Calvin cycle is carboxylation. Here, CO₂ is fixed into a stable organic intermediate. RuBP is a 5-C compound. It is carboxylated by utilising CO₂ and then C-C bond cleavage results in the formation of 2 molecules of 3-PGA.

The reaction catalysed by RuBisCO is as follows:

RuBP (5C) + CO₂ + H₂O → 2 3-PGA (3C)

The reaction involves enolisation of RuBP followed by carboxylation, which leads to the formation of an intermediate 3-keto-2′-carboxyarabinitol-1,5-bisphosphate. It is followed by hydration, and then subsequent cleavage of the bond between two carbons to give rise to 2 molecules of 3-phosphoglycerate (3-PGA). The 3-PGA thus formed is utilised in the formation of glucose and other carbohydrates in the subsequent steps. In C₃ plants, this process occurs in the mesophyll cells. In the C₄ pathway, the Clavin cycle occurs in the bundle sheath cells. The bundle sheath cells are rich in RuBisCO. This is an adaptation to reduce photorespiration in C₄ plants.

Photorespiration

RuBisCO also has an affinity for oxygen and it oxygenates RuBP in the presence of oxygen. Photorespiration utilises ATP, hence, leads to the wasting of some energy produced in photosynthesis. When RuBisCO binds to O₂ it converts RuBP to one molecule of phosphoglycerate (3C) and phosphoglycolate (2 Carbon) each. It is a waste process, it neither generates ATP nor sugar.

Characteristics of RUBISCO

• RuBisCO is found in photosynthetic protists, algae, plants  and some autotrophic bacteria like cyanobacteria and proteobacteria.
• The most plentiful protein in the biosphere is RuBisCO. It accounts for approximately 30% and 50% of soluble solid leaf nutrients in C4 and C3 plants, respectively.
• RuBisCO is found in mesophyll cells in C3 plants but not in bunch sheath cells in C4 plants.
• It is a large and complex protein with small and large chains. The molecular weight is approximately 540,000 Da.
• In general, there are eight large chains which form four dimers and eight small chains. The large chain contains an active location for the substrate. Large subunits are the only ones found in some bacteria and dinoflagellates.

Functions of RUBISCO

• RuBisCO’s primary function is photosynthesis and photorespiration.
• It catalyses the very first step in the Calvin cycle or C3 pathway, namely the carboxylation of RuBP. This results in formation of two 3-PGA molecules.
• RuBisCO has oxygen affinity, so it unites to some O, during the photorespiration process. It converts RuBP to one molecule of each phosphoglycerate and phosphoglycolate.
• Because RuBisCO has a much stronger affinity for CO2 when compared to O2, photosynthesis is favoured over photorespiration.

Q.3 DESCRIBE PLANT RESPIRATION?

Respiration in Plants
Types of Respiration - Aerobic and Anaerobic Respiration

All living organisms, containing plants, get the energy necessary for their survival from a series of chemical reactions termed respiration. The process of respiration needs glucose to start the reactions which are changed into energy and later produce carbon dioxide and water as by-products

Plant Respiration

The method by which cells get chemical energy by the consumption of oxygen and the liberating of carbon dioxide is called respiration. In order to carry on respiration, plant cells require oxygen and a means of disposing of carbon dioxide just as animal cells do. In plants, every part such as root, stem executes respiration as plants do not possess any particular organs like animals for the exchange of gases.The method of respiration is written as:

Oxygen + Glucose → Water + Carbon Dioxide with Energy

Conclude the same from the equation above as well that respiration uses oxygen and to produce carbon dioxide.

Role of Air Temperature
Plant respiration happens 24 hours a day, but night respiration is more obvious as the photosynthesis process finishes. During the night, it is very vital that the temperature is much cooler as compared to the daytime because plants can undergo stress. Imagine a marathon runner. The runner breathes at higher rates than an individual standing still; so, a runner’s amount of respiration is greater and the temperature of the body rises. The same principle relates to plants, as the temperature at night rises, the respiration rate increases, and similar temperature increases. This action would result in flower damage and also in plant poor growth.

Respiration in Roots
In plants, respiration occurs with the help of roots. In soil oxygenated air is already present in spaces between soil particles. This oxygen is then absorbed into the roots with the help of root hair present on the roots. The hairs of the roots are in straight contact with them. In fact, root hair is a lateral tubular outgrowth of the external epidermal cells of a root. The oxygen present among the soil particles diffuses into the root hairs. From root hairs, oxygen is transported to all the parts of roots for respiration. During the respiration process, oxygen is transformed into carbon dioxide gas which is spread in the opposite direction i.e. out of the roots by the same root hairs which complete the respiration process of roots.

If a potted plant is watered over for a long time, then the plant can ultimately die. This is due to too much water exorcizing all the air from in between the soil particles. Because of this, oxygen is not free to the roots for aerobic respiration. In these states, the roots of plants respire anaerobically making alcohol. This can kill the plant. Germinating seeds during the early stage respire anaerobically as they have a seed coat that does not permit the oxygen to enter through it.

Respiration in plants occurs throughout the day and night thereby carbon dioxide is formed. Though, during the day, the total of carbon dioxide CO2 released is insignificant compared to the volume of oxygen made as a result of photosynthesis. Therefore, one should not sleep underneath a tree at night.

Respiration in Stems
In the plants taking herbaceous stem exchange of gases occurs through stomata and the carbon dioxide CO2 formed during the process that gets diffused into the air with the help of stomata only. While in the plants having hard and woody stems the exchange of gases occurs through lenticels. Lenticels are usually loosely packed dead cells that are present as tiny pores on the bark of woody plants. These allow oxygen to pass to the intercellular spaces of the inside of tissues and carbon dioxide (CO2) to be liberated into the atmosphere by the phenomena of diffusion which completes the process of respiration in stems.

Respiration in Leaves
In leaves, the exchange of respiratory gases occurs through very small pores called stomata. The stomata are present in big numbers on the lower side of the leaves of the plant. Every stoma has a tiny pore at its centre which is enclosed and regulated by two kidney-shaped cells known as guard cells. When the stoma opens the exchange of gases occurs between the atmosphere and interior of the leaf by the method of diffusion and that completes the process of respiration in leaves

Types of Respiration

There are two kinds of respiration that we categorize on the basis of the absence or presence of oxygen:

1_Aerobic Respiration

The respiration that occurs in the presence of oxygen is named aerobic respiration due to ‘air’ which has oxygen. Aerobic respiration contains the utilization of oxygen for the breaking of chemical bonds in glucose to liberate energy in high volumes. It is the central source of energy for plants. Animals and plants that use oxygen for respiration are aerobes. Most animals have aerobic respiration.

C₆H₁₂O₆ +6O₂ ⟶ 6CO₂ + 6H₂O + Energy

All the organisms that gain energy by aerobic respiration cannot exist without oxygen. This is due to no oxygen there; they cannot get energy from the food which they consume. Aerobic respiration takes more energy because a complete breaking of glucose takes place during respiration with the use of oxygen.

2_.Anaerobic Respiration

The respiration that occurs in the absence of oxygen is known as anaerobic respiration. In this process, the incomplete oxidation of food substances is being made by carbon dioxide CO2, and alcohol(OH). Besides this other organic matter such as citric acid, oxalic acid, lactic acid, etc are also produced.

This process is also called intramolecular respiration. Anaerobic respiration occurs in organisms like yeast, certain bacteria, and parasitic worms. The animals and plants that can exist and gain energy even in the lack of oxygen are called Anaerobic.

Glucose ⟶ Alcohol + CO2 + (Energy)

Yeast is known to be a single-celled fungus. In yeast, a single cell signifies the whole organism. A very low volume of energy is liberated in this process. Yeast respires anaerobically and all through this process, yeast transforms glucose into alcohol. So it is used to make alcohol, bread, etc. Anaerobic respiration yields much less energy due to the only partial breakdown of glucose happening in anaerobic respiration in the absence of oxygen. All the organisms which gain energy by anaerobic respiration can exist without oxygen. For instance; yeast is an organism that can exist without the oxygen of air as it obtains energy by the method of anaerobic respiration. Yeast can live in the absence of oxygen.

Q.4 EXPLAIN C4 PHOTOSYNTHESIS IN DETAIL?

The chemical process of photosynthesis which takes place independent of light is referred to as dark reaction. It occurs in the stroma of the chloroplast. This dark reaction is enzymatic purely, and compared to the light reaction, is slower. Dark reactions also take place when light is present. The sugars in the dark reactions are synthesized from carbon dioxide. The energy-deprived CO₂ is fixed to energy-rich carbohydrates utilizing energy-rich compound, ATP and the assimilatory power, the NADPH₂ of light reaction. The process is referred to as carbon assimilation or carbon fixation.

As Blackman illustrated the existence of a dark reaction. This reaction was referred to as Blackman’s reaction. There are two types of cyclic reactions occurring in the dark reaction –

• Calvin cycle or C3 cycle
• Hatch and Slack pathway or C4 cycle

Hatch and Slack Pathway

M. D. Hatch and C. R. Slack first outlined this metabolic pathway in depth. In this, the carbon dioxide is added first to the phosphoenolpyruvate by the action of the enzyme PEP carboxylase. Thus, producing the four-carbon compound in the mesophyll cells, this is transported later on to the bundle sheath cells to release the carbon dioxide to be used in the Calvin cycle. In 1966, Hatch and Slack discovered the C4 cycle, hence the name. It is also referred to as the ß carboxylation pathway and co-operative photosynthesis. The 4-carbon oxaloacetic acid is the first stable compound of the Hatch and Slack cycle, hence is called the C4 cycle.C4 plants are plants possessing a C4 cycle. Such plants are inclusive of dicots and monocots, the C4 cycle is evident in the Chenopodiaceae, Gramineae and Cyperaceae family.

Hatch and Slack Pathway In C4 Plants

To fix carbon dioxide, this pathway is the alternate to the C3 cycle. Here, the first formed stable compound – oxaloacetic acid is a 4 carbon compound, hence the name C4 cycle. This pathway is a common sight in several grasses, maize, sugarcane, amaranthus, sorghum. The C4 plants depict a different kind of leaf anatomy (Kranz anatomy).

The chloroplasts are dimorphic and in the leaves, vascular bundles are wrapped by a bundle sheath of larger parenchymatous cells. Such bundle sheath cells possess chloroplasts, which are larger, containing starch grains, lacking grana while the chloroplasts in the mesophyll cells always possess grana and are smaller. The bundle sheath cells appear as a wreath or a ring while cells are larger. This characteristic leaf anatomy of the C4 plants is referred to as Kranz Anatomy. In German, Kranz corresponds to the wreath, hence the name Kranz Anatomy.

The C4 Cycle depicts two carboxylation reactions occurring in the chloroplasts of the mesophyll cells and others in the chloroplast of the bundle sheath cells. The Hatch and Slack Cycle involves four steps –

• Carboxylation
• Breakdown
• Splitting
• Phosphorylation
•  

Carboxylation

Occurs in the chloroplasts of the mesophyll cells. A 3-carbon compound, Phosphoenolpyruvate, collects carbon dioxide and in the presence of water, transforms to 4 carbon oxaloacetate. The enzyme phosphoenolpyruvate carboxylase catalyzes the reaction.

Breakdown

Readily, oxaloacetate disintegrates into 4 carbon malate and aspartate. The enzyme involved in the reaction is transaminase and malate dehydrogenase. The compounds formed diffuse into the sheath cells from the mesophyll cells.

Splitting

The malate and aspartate in the sheath cells enzymatically split to produce free carbon dioxide and 3-carbon pyruvate. The carbon dioxide is made use of Calvin’s cycle in the sheath cells. The second carboxylation takes place in the chloroplasts of the bundle sheath cells. The carbon dioxide is accepted by the 5-carbon compound ribulose diphosphate with the activity of the carboxy dismutase enzyme, finally producing 3 phosphoglyceric acid. For the formation of sugars, some of the 3 phosphoglyceric acid is used and the remaining regenerates ribulose diphosphate.

Phosphorylation

The pyruvate molecules are moved to the chloroplasts of the mesophyll cells wherein, in the presence of ATP, it is phosphorylated for the regeneration of phosphoenolpyruvate. Pyruvate phosphokinase catalyzes the reaction and phosphoenolpyruvate is regenerated.

In this pathway, the C3 and C4 cycles of the carboxylation are associated as a result of the Kranz anatomy of the leaves. Compared to C3 plants, the C4 plants are more efficient in photosynthesis. Phosphoenolpyruvate carboxylase enzyme of the C4 cycle is seen to possess more affinity for carbon dioxide compared to ribulose diphosphate carboxylase of the C3 cycle when it comes to fixing molecular carbon dioxide in the organic compound at the time of carboxylation.

Q.5 Explain modern concept of electron transport chain

Definition -The electron transport chain is a cluster of proteins that transfer electrons through a membrane within mitochondria to form a gradient of protons that drives the creation of adenosine triphosphate (ATP). ATP is used by the cell as the energy for metabolic processes for cellular functions.

During the process, a proton gradient is created when the protons are pumped from the mitochondrial matrix into the intermembrane space of the cell, which also helps in driving ATP production. Often, the use of a proton gradient is referred to as the chemiosmotic mechanism that drives ATP synthesis since it relies on a higher concentration of protons to generate “proton motive force”. The amount of ATP created is directly proportional to the number of protons that are pumped across the inner mitochondrial membrane.

The electron transport chain involves a series of redox reactions that relies on protein complexes to transfer electrons from a donor molecule to an acceptor molecule. As a result of these reactions, the proton gradient is produced, enabling mechanical work to be converted into chemical energy, allowing ATP synthesis. The complexes are embedded in the inner mitochondrial membrane called the cristae in eukaryotes. Enclosed by the inner mitochondrial membrane is the matrix, which is where necessary enzymes such as pyruvate dehydrogenase and pyruvate carboxylase are located. The process can also be found in photosynthetic eukaryotes in the thylakoid membrane of chloroplasts and in prokaryotes, but with modifications.

By-products from other cycles and processes, like the citric acid cycle, amino acid oxidation, and fatty acid oxidation, are used in the electron transport chain. As seen in the overall redox reaction,

2 H⁺ + 2 E⁺ + ½ O₂ → H₂O + ENERGY

Energy is released in an exothermic reaction when electrons are passed through the complexes; three molecules of ATP are created. Phosphate located in the matrix is imported via the proton gradient, which is used to create more ATP. The process of generating more ATP via the phosphorylation of ADP is referred to oxidative phosphorylation since the energy of hydrogen oxygenation is used throughout the electron transport chain. The ATP generated from this reaction go on to power most cellular reactions necessary for life.

Steps of the Electron Transport Chain

In the electron transfer chain, electrons move along a series of proteins to generate an expulsion type force to move hydrogen ions, or protons, across the mitochondrial membrane. The electrons begin their reactions in Complex I, continuing onto Complex II, traversed to Complex III and cytochrome c via coenzyme Q, and then finally to Complex IV. The complexes themselves are complex-structured proteins embedded in the phospholipid membrane. They are combined with a metal ion, such as iron, to help with proton expulsion into the intermembrane space as well as other functions. The complexes also undergo conformational changes to allow openings for the transmembrane movement of protons.

These four complexes actively transfer electrons from an organic metabolite, such as glucose. When the metabolite breaks down, two electrons and a hydrogen ion are released and then picked up by the coenzyme NAD⁺ to become NADH, releasing a hydrogen ion into the cytosol.

The NADH now has two electrons passing them onto a more mobile molecule, ubiquinone (Q), in the first protein complex (Complex I). Complex I, also known as NADH dehydrogenase, pumps four hydrogen ions from the matrix into the intermembrane space, establishing the proton gradient. In the next protein, Complex II or succinate dehydrogenase, another electron carrier and coenzyme, succinate is oxidized into fumarate, causing FAD (flavin-adenine dinucleotide) to be reduced to FADH₂. The transport molecule, FADH₂ is then reoxidized, donating electrons to Q (becoming QH₂), while releasing another hydrogen ion into the cytosol. While Complex II does not directly contribute to the proton gradient, it serves as another source for electrons.

Complex III, or cytochrome c reductase, is where the Q cycle takes place. There is an interaction between Q and cytochromes, which are molecules composed of iron, to continue the transfer of electrons. During the Q cycle, the ubiquinol (QH₂) previously produced donates electrons to ISP and cytochrome b becoming ubiquinone. ISP and cytochrome b are proteins that are located in the matrix that then transfers the electron it received from ubiquinol to cytochrome c1. Cytochrome c1 then transfers it to cytochrome c, which moves the electrons to the last complex. (Note: Unlike ubiquinone (Q), cytochrome c can only carry one electron at a time). Ubiquinone then gets reduced again to QH₂, restarting the cycle. In the process, another hydrogen ion is released into the cytosol to further create the proton gradient.

The cytochromes then extend into Complex IV, or cytochrome c oxidase. Electrons are transferred one at a time into the complex from cytochrome c. The electrons, in addition to hydrogen and oxygen, then react to form water in an irreversible reaction. This is the last complex that translocates four protons across the membrane to create the proton gradient that develops ATP at the end.

As the proton gradient is established, F₁F₀ ATP synthase, sometimes referred to as Complex V, generates the ATP. The complex is composed of several subunits that bind to the protons released in prior reactions. As the protein rotates, protons are brought back into the mitochondrial matrix, allowing ADP to bind to free phosphate to produce ATP. For every full turn of the protein, three ATP is produced, concluding the electron transport chain.